The present invention relates to a novel class of organic molecules capable of inhibiting the enzyme C-proteinase, pharmaceutical compositions comprising the C-proteinase inhibitory compounds and methods of using the compounds and compositions to regulate, modulate and/or inhibit collagen production and/or maturation as a therapeutic approach towards the treatment or prevention of myriad diseases related to, or associated with, unregulated collagen production.
At present, nineteen different types of collagens have been identified. These collagens, which include fibrillar collagen types I, II and III, are synthesized as procollagen precursor molecules which contain peptide extensions at both their amino- and carboxy-termini. These peptide extensions, referred to as xe2x80x9cpro-regions,xe2x80x9d are designated as N- and C-propeptides, respectively.
The pro-regions are typically cleaved upon secretion of the procollagen triple helical precursor molecule from the cell to yield a mature triple helical collagen molecule. Upon cleavage, the xe2x80x9cmaturexe2x80x9d collagen molecule is capable of association, for example, into highly structured collagen fibers. See e.g., Fessler and Fessler, 1978, Annu. Rev. Biochem. 47:129-162; Bornstein and Traub, 1979, in: The Proteins (eds. Neurath, H. and Hill, R. H.), Academic Press, New York, pp. 412-632; Kivirikko et al., 1984, in: Extracellular Matrix Biochemistry (eds. Piez, K. A. and Reddi, A. H.), Elsevier Science Publishing Co., Inc., New York, pp. 83-118; Prockop and Kivirikko, 1984, N. Engl. J. Med. 311:376-383; Kuhn, 1987, in: Structure and Function of Collagen Types (eds. Mayne, R. and Burgeson, R. E.), Academic Press, Inc., Orlando, Fla., pp. 1-42.
A variety of critical diseases have been linked to inappropriate or unregulated collagen production and/or maturation. These diseases include pathological fibrosis or scarring (including endocardial sclerosis), idiopathic interstitial fibrosis, interstitial pulmonary fibrosis, perimuscular fibrosis, Symmers"" fibrosis, pericentral fibrosis, hepatitis, dermatofibroma, binary cirrhosis, alcoholic cirrhosis, acute pulmonary fibrosis, idiopathic pulmonary fibrosis, acute respiratory distress syndrome, kidney fibrosis/glomerulonephritis, kidney fibrosis/diabetic nephropathy, scleroderma/systemic, scleroderma/local, keloids, hypertrophic scars, severe joint adhesions/arthritis, myelofibrosis, corneal scarring, cystic fibrosis, muscular dystrophy (duchenne""s), cardiac fibrosis, muscular fibrosis/retinal separation, esophageal stricture and payronles disease. Further fibrotic disorders may be induced or initiated by surgery, including scar revision/plastic surgeries, glaucoma, cataract fibrosis, corneal scarring, joint adhesions, graft vs. host disease, tendon surgery, nerve entrapment, dupuytren""s contracture, OB/GYN adhesions/fibrosis, pelvic adhesions, peridural fibrosis, restenosis. One strategy for the treatment of these diseases is to inhibit the pathological overproduction of collagen. Thus, identification and isolation of molecules which control, inhibit and/or modulate the production of collagen are of major medical interest.
Recent evidence suggests that C-proteinase is the essential key enzyme that catalyzes the cleavage of the C-propeptide of, for example, fibrillar collagens, including type I, type II, and type III collagen. See e.g. Prockep et al., 1998, Matrix Biol. 16:399-408; Lee et al., 1997, J. Biol. Chem. 272:19059-19066; Suzuk et al., 1996, Development 122:3587-3595.
C-proteinase was first observed in the culture media of human and mouse fibroblasts (Goldberg et al., 1975, Cell 4:45-50; Kessler and Goldberg, 1978, Anal. Biochem. 86:463-469), and chick tendon fibroblasts (Duskin et al., 1978, Arch. Biochem. Biophys. 185:326-332; Leung et al., 1979, J. Biol. Chem. 254:224-232). An acidic proteinase which removes the C-terminal propeptides from type I procollagen has also been identified. Davidson et al., 1979, Eur. J. Biochem. 100:551.
A partially purified protein having C-proteinase activity was obtained from chick calvaria in 1982. Njieha et al., 1982, Biochemistry 23:757-764. In 1985, chicken C-proteinase was isolated, purified and characterized from conditioned media of chick embryo tendons. Hojima et al., 1985, J. Biol. Chem. 260:15996-16003. Murine C-proteinase has been subsequently purified from media of cultured mouse fibroblasts. Kessler et al., 1986, Collagen Relat. Res. 6:249-266; Kessler and Adar, 1989, Eur. J. Biochem. 186:115-121. Finally, the cDNA encoding human C-proteinase has been identified (see, e.g., Takahara et al., 1994, J. Biol. Chem. 269:26280-26285; Li et al., 1996, Proc. Natl. Acad. Sci. USA 93:5127-5130; Kessler et al., 1996, Science 271:360-362.
Experiments conducted with purified forms of chick and mouse C-proteinases indicate that C-proteinase is instrumental in the formation of functional collagen fibers. Fertala et al., 1994, J. Biol. Chem. 269:11584. As a consequence of its critical role in collagen production and maturation, scientists have sought to identify compounds that inhibit C-proteinase. See e.g., Hojima et al., supra. Compounds identified to date include metal chelators (e.g., EDTA, phenanthroline, EGTA, basic amino acids (e.g., lysine and arginine), peptides (e.g., chymostatin, pepstatin A, and concanavalin A), proteins (e.g., xcex12-macroglobulin, ovostatin, and fetal bovine serum), metals ions (e.g., Zn2+, Cu2+, and Cd2+), reducing agents (e.g., dithiothreitol), detergents (e.g., sodium dodecyl sulfate (SDS)) and certain salts and buffers (e.g., phosphate, ammonium sulfate, sodium chloride and tris hydrochloride). In contrast, microbial inhibitors such as leupeptin, phosphoramidon, antipain, bestatin, elastinal, and amastatin are considered to have weak or no effect on the activity of C-proteinase. For references discussing the various C-proteinase inhibitors identified to date, see Leung et al., supra; Ryhxc3xa4nen et al., 1982, Arch. Biochem. Biophys. 215:230-236; W097/05865; and the references cited therein.
C-proteinase belongs to the matrix metalloproteinase (MMP) superfamily of zinc endopeptidases which are involved in tissue remodeling. Members of the MMP family include MMP-1 (human collagenase), MMP-2 (gelatinase), and MMP-9 (human gelatinase B). See e.g. W098/34918; Krumme et al., 1998, FEBS Lett. 436:209-212. The MMPs are characterized by an active site zinc ion that plays an essential role in the enzymatic activity of MMPs. Rational drug discovery efforts, involving the inhibition of MMPs, have focused on inhibitor classes that contain a functional group that can coordinate the zinc ion and thereby inactivate the target MMP. See e.g. Krumme et al., supra. One such inhibitor class are hydroxamic acids. As revealed by the x-ray crystal structure determination of hydroxamic acid:MMP cocrystals, the hydroxamic acid coordinates the active site zinc in a bidentate manner via the hydroxyl and carbonyl oxygens of the hydroxamic group. See Grams et al., 1995, Biochem. 34:14012-14020; Bode et al., 1994, EMBO J., 13:1263-1269. Despite their potent affinity as zinc coordinators, hydroxamic acids demonstrate a considerable degree of specificity within the MMP family. Thus a potent inhibitor of MMP-1 (human collagenase) may have only minimal potency against another MMP such as C-proteinase. Thus the development of potent hydroxamic acid inhibitors against a particular MMP requires considerable research effort and experimentation.
In view of its essential role in the formation and maturation of collagen, C-proteinase provides an ideal therapeutic target towards the treatment or prevention of disorders related to, or associated with, unregulated collagen production or maturation. Accordingly, there remains a need in the art for compounds that are specific and potent inhibitors of C-proteinase, especially C-proteinase inhibitory compounds which provide clinically relevant benefits in the treatment or prevention of diseases associated with unregulated collagen production and/or maturation.
In one aspect, the present invention provides a novel class of organic molecules that are potent and/or selective inhibitors of C-proteinase. As a consequence of this activity, the compounds of the invention are capable of modulating, regulating or inhibiting collagen production or maturation by affecting C-proteinase activity.
The compounds of the invention are generally substituted hydantoin compounds according to structural formulae (Ia) and (Iaxe2x80x2): 
or pharmaceutically acceptable salts thereof, wherein:
m is an integer from 0 to 3 when X is NR19 or an integer from 1 to 3 when X is S or O;
p is an integer from 1 to 4 when X is NR19 or an integer from 1 to 3 when X is S or O;
n is and integer from 1 to 3;
X is NR19, S or O;
R19 is selected from the group consisting of hydrogen, (C1-C5) alkyl, and (C1-C5) substituted alkyl;
R1 and R2 are each independently selected from the group consisting of (C1-C10) alkyl, (C1-C10) substituted alkyl, (C3-C10) alkenyl, (C3-C10) substituted alkenyl, (C3-C10) alkynyl, (C3-C10) substituted alkynyl, (C3-C20) cycloalkyl, (C3-C20) substituted cycloalkyl, 3-20 membered heterocycloalkyl, 3-20 membered substituted heterocycloalkyl, (C5-C20) aryl, (C5-C20) aryl independently substituted with one or more Y1, alkaryl, alkaryl independently substituted with one or more Y1, 5-20 membered heteroaryl, 5-20 membered heteroaryl independently substituted with one or more Y1, 6-26 membered alk-heteroaryl, and 6-26 membered alk-heteroaryl independently substituted with one or more Y1;
R20 and R21 are each independently selected from the group consisting of hydrogen, (C1-C10) alkyl, (C6-C26) alkaryl, and 6-26 membered alk-heteroaryl; and
each Y1 is independently selected from the group consisting of -halogen, -trihalomethyl, xe2x80x94R, xe2x80x94C(O)OR, xe2x80x94CN, xe2x80x94C(O)xe2x80x94NRxe2x80x94OR, xe2x80x94C(NRR)xe2x95x90Nxe2x80x94OR, xe2x80x94C(O)xe2x80x94R, xe2x80x94C(O)NRR, xe2x80x94C(S)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94NRR, xe2x80x94NO2, xe2x80x94N3, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94OR, xe2x80x94P(O) (OH) (NRR), xe2x80x94P(O) (OH)21, xe2x80x94SO2R, xe2x80x94S(O)xe2x80x94R, xe2x80x94SO3H, xe2x80x94SR, xe2x80x94SO2xe2x80x94NRR, xe2x80x94OCF3 and -heteroaryl; where each R is independently selected from the group consisting of, hydrogen, (C1-C8) alkyl, (C3-C8) alkenyl, (C5-C20) aryl, (C6-C26) alkaryl, 6-20 membered heteroaryl and 6-26-membered alk-heteroaryl;
with the proviso that the sum of m and p is 1, 2, 3 or 4.
In a preferred embodiment, the compounds of the invention iS are generally substituted hydantoin compounds having the structural formula (Ib): 
or pharmaceutically acceptable salts thereof, wherein:
a is an integer from 2 to 5; and
R1, R2, and R20 are as defined for formula (Ia).
In another aspect, the present invention provides pharmaceutical compositions comprising a therapeutically effective amount of one or more of the above-described compounds and a pharmaceutically acceptable carrier, diluent or excipient. Such a composition can be used in the methods of the invention to inhibit, regulate or modulate the production or maturation of collagen by inhibiting C-proteinase activity and to treat or prevent a variety of collagen-related disorders.
In yet another aspect, the invention provides methods of inhibiting C-proteinase activity. The method involves contacting the enzyme C-proteinase, or an active fragment or derivative thereof, with an amount of a compound or composition according to the invention effective to block collagen production. Methods of inhibiting C-proteinase in vivo can be used to inhibit, regulate or modulate collagen production or maturation as a therapeutic approach towards the treatment or prevention of disorders related to, or associated with, unregulated collagen production or maturation.
In a final aspect, the present invention provides methods for the treatment or prevention of disorders related to, or associated with, inappropriate or unregulated collagen production or maturation. The method involves administering if to an animal subject, including a human, an amount of a compound according to the invention, or a pharmaceutical composition thereof, effective to treat or prevent the particular collagen-related disorder.
Disorders which can be treated or prevented according to the methods of the invention include, but are not limited to, rheumatoid arthritis, scleroderma, pathological fibrosis or scarring.
As used herein, the following terms shall have the following meanings:
xe2x80x9cC-proteinasexe2x80x9d: refers to an enzyme capable of processing collagen molecules, derivatives or fragments of collagen molecules or precursors of collagen molecules, collagen derivatives or collagen fragments by cleaving the amino acid sequence -Ala↓Asp-Asp-, -Gly↓Asp-Glu- and/or -Ala↓Asp-Gln- at the position marked with xe2x80x9c↓xe2x80x9d. The term xe2x80x9cC-proteinasexe2x80x9d includes human C-proteinase as well as derivatives, analogs, fragments and variants thereof capable of processing collagen molecules as described above.
xe2x80x9cAlkylxe2x80x9d: refers to a saturated branched, straight chain or cyclic hydrocarbon radical. Typical alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, cyclopropyl, butyl, isobutyl, t-butyl, cyclobutyl, pentyl, isopentyl, cyclopentyl, hexyl, cyclohexyl and the like. In preferred embodiments, the alkyl groups are (C1-C8) alkyl, more preferably (C1-C6) alkyl and most preferably (C1-C3) alkyl.
xe2x80x9cSubstituted Alkylxe2x80x9d: refers to an alkyl radical that has been substituted with one or more Q. Typical substituents Q include, but are not limited to, xe2x80x94X, xe2x80x94OR, xe2x80x94NRR, xe2x80x94N3, xe2x80x94CN, and xe2x80x94CX3; where each R is independently hydrogen, alkyl or substituted alkyl, and X is halogen.
xe2x80x9cAlkenylxe2x80x9d: refers to an unsaturated branched, straight chain or cyclic hydrocarbon radical having at least one carbon-carbon double bond. The radical may be in either the cis or trans conformation about the double bond(s). Typical alkenyl groups include, but are not limited to, ethenyl, propenyl, isopropenyl, isopropylidene, butenyl, butenylidene, isobutenyl, tert-butenyl, cyclobutenyl, pentenyl, isopentenyl, cyclopentenyl, hexenyl, cyclohexenyl and the like. In preferred embodiments, the alkenyl group is (C2-C8) alkenyl, more preferably (C2-C6) alkenyl and most preferably (C3-C4) alkenyl.
xe2x80x9cSubstituted Alkenylxe2x80x9d: refers to an alkenyl radical that has been substituted with one or more Q. Typical Q include, but are not limited to, xe2x80x94X, xe2x80x94OR, xe2x80x94NRR, xe2x80x94N3, xe2x80x94CN, and xe2x80x94CX3; where each R is independently hydrogen, alkyl or substituted alkyl, and X is halogen.
xe2x80x9cAlkynylxe2x80x9d: refers to an unsaturated branched, straight-chain or cyclic alkyl group having at least one carbon-carbon triple bond. Typical alkynyl groups include, but are not limited to, ethynyl; propynyls such as prop-1-yn-1-yl , prop-2-yn-1-yl, etc.; butynyls such as but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl , etc.; and the like. In preferred embodiments, the alkynyl group is (C3-C6) alkynyl.
xe2x80x9cSubstituted Alkynylxe2x80x9d: refers to an alkynyl radical that has been substituted with one or more Q. Typical Q include, but are not limited to, xe2x80x94X, xe2x80x94OR, xe2x80x94NRR, xe2x80x94N3, xe2x80x94CN, and xe2x80x94CX3; where each R is independently hydrogen, alkyl or substituted alkyl, and X is halogen.
xe2x80x9cCycloalkylxe2x80x9d: refers to a cyclic or polycyclic saturated or unsaturated hydrocarbon radical. Typical cycloalkyl groups include, but are not limited to, cyclopropanyl, cyclobutanyl, cyclopentanyl, cyclohexanyl and higher cycloalkyls, adamantyl, cubanyl, prismanyl and higher polycylicalkyls, etc. In preferred embodiments, the cycloalkyl is (C3-C10) cycloalkyl. Particularly preferred cycloalkyls are cyclohexanyl and adamantyl.
xe2x80x9cSubstituted Cycloalkylxe2x80x9d: refers to a cycloalkyl or radical wherein one or more hydrogen atoms are each independently replaced with another substituent. Typical substituents include, but are not limited to, (C1-C8) alkyl, (C3-C8) alkenyl, (C3-C8) alkynyl, (C5-C20) aryl, (C6-C26) alkaryl, xe2x80x94OR, xe2x80x94SR, xe2x80x94NRR, xe2x80x94CN, xe2x80x94NO2, xe2x80x94C(O)R, xe2x80x94C(O)OR, xe2x80x94C(O)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94C(O)NROR, xe2x80x94C(NRR)xe2x95x90NOR, xe2x80x94NRxe2x80x94C(O)R, -tetrazol-5-yl, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, -halogen, and -trihalomethyl; where each R is independently xe2x80x94H, (C1-C8) alkyl, (C3-C8) alkenyl, (C3-C8) alkynyl, (C5-C20) aryl, and (C6-C26) alkaryl as defined herein.
xe2x80x9cHeterocycloalkylxe2x80x9d: refers to a cycloalkyl moiety wherein one of the ring carbon atoms is replaced with another atom such as N, P, O, S, As, Ge, Se, Si, Te, etc. Typical heterocycloalkyls include, but are not limited to, imidazolidyl, piperazyl, piperidyl, pyrazolidyl, pyrrolidyl, quinuclidyl, etc. In preferred embodiments, the cycloheteroalkyl is 5-10 membered. Particularly preferred cycloheteroalkyls are morpholino, tetrahydrofuryl, and pyrrolidyl.
xe2x80x9cSubstituted Heterocycloalkylxe2x80x9d: refers to a cycloheteroalkyl radical wherein one or more hydrogen atoms are each independently replaced with another substituent. Typical substituents include, but are not limited to, (C1-C8) alkyl, (C3-C8) alkenyl, (C3-C8) alkynyl, (C5-C20) aryl, (C6-C26) alkaryl, 5-20 membered heteroaryl, 6-26 membered alk-heteroaryl, xe2x80x94OR, xe2x80x94SR, xe2x80x94NRR, xe2x80x94CN, xe2x80x94NO2, xe2x80x94C(O)R, xe2x80x94C(O)OR, xe2x80x94C(O)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94C(O)NROR, xe2x80x94C(NRR)xe2x95x90NOR, xe2x80x94NRxe2x80x94C(O)R, -tetrazol-5-yl, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, -halogen and -trihalomethyl; where each R is independently xe2x80x94H, (C1-C8) alkyl, (C2-C8) alkenyl, (C2-C8) alkynyl, (C5-C20) aryl, (C6-C26) alkaryl, 5-20 membered heteroaryl, and 6-26 membered alk-heteroaryl as defined herein.
xe2x80x9cArylxe2x80x9d: refers to an unsaturated cyclic hydrocarbon radical having a conjugated Π electron system. Specifically included within the definition of aryls are fused ring systems in which one or more of the rings are aromatic and one or more of the rings are saturated or unsaturated, such as, for example, indane, indene, phenalene, etc. Typical aryl groups include, but are not limited to, penta-2,4-dienyl, phenyl, naphthyl, aceanthrylyl, acenaphthyl, anthracyl, azulenyl, chrysenyl, indacenyl, indanyl, ovalenyl, perylenyl, phenanthrenyl, phenalenyl, picenyl, pyrenyl, pyranthrenyl, rubicenyl and the like. In preferred embodiments, the aryl group is (C5-C20) aryl, more preferably (C5-C10) aryl and most preferably phenyl.
xe2x80x9cAlkarylxe2x80x9d: refers to a straight-chain or branched (C1-C8) alkyl, (C2-C8) alkenyl or (C2-C8) alkynyl group wherein one of the hydrogen atoms bonded to a terminal carbon is replaced with a (C5-C20) aryl moiety. Typical alkaryl groups include, but are not limited to, benzyl, benzylidene, benzylidyne, benzenobenzyl, naphthalenobenzyl and the like. In preferred embodiments, the alkaryl group is (C6-C26) alkaryl, i.e., the alkyl, alkenyl or alkynyl moiety of the alkaryl group is (C1-C6) and the aryl moiety is (C5-C20) In particularly preferred embodiments the alkaryl group is (C6-C13), i.e., the alkyl, alkenyl or alkynyl moiety of the alkaryl group is (C1-C3) and the aryl moiety is (C5-C10).
xe2x80x9cHeteroarylxe2x80x9d: refers to an aryl moiety wherein one or more carbon atoms has been replaced with another atom, such as N, P, O, S, As, Ge, Se, Si, Te, etc. Typical heteroaryl groups include, but are not limited to acridarsine, acridine, arsanthridine, arsindole, arsindoline, benzodioxole, benzothiadiazole, carbazole, xcex2-carboline, chromane, chromene, cinnoline, furan, imidazole, indazole, indole, isoindole, indolizine, isoarsindole, isoarsinoline, isobenzofuran, isochromane, isochromene, isoindole, isophosphoindole, isophosphinoline, isoquinoline, isothiazole, isoxazole, naphthyridine, perimidine, phenanthridine, phenanthroline, phenazine, phosphoindole, phosphinoline, phthalazine, piazthiole, pteridine, purine, pyran, pyrazine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole, pyrrolizine, quinazoline, quinoline, quinolizine, quinoxaline, selenophene, tellurophene, thiazopyrrolizine, thiophene and xanthene. In preferred embodiments, the heteroaryl group is a 5-20 membered heteroaryl, with 5-10 membered heteroaryl being particularly preferred.
xe2x80x9cAlk-heteroarylxe2x80x9d: refers to a straight-chain or branched (C1-C8) alkyl, (C2-C8) alkenyl or (C2-C8) alkynyl group wherein one of the hydrogen atoms bonded to a terminal carbon atom is replaced with a heteroaryl moiety. In preferred embodiments, the alk-heteroaryl group is a 6-26 membered alk-heteroaryl, i.e., the alkyl, alkenyl or alkynyl moiety of the alk-heteroaryl is (C1-C6) and the heteroaryl moiety is a 5-20-membered heteroaryl. In particularly preferred embodiments, the alk-heteroaryl is a 6-13 membered alk-heteroaryl, i.e., the alkyl, alkenyl or alkynyl moiety is (C1-C3) and the heteroaryl moiety is a 5-10 membered heteroaryl.
The present invention provides a novel class of organic compounds capable of inhibiting the enzyme C-proteinase, pharmaceutical compositions comprising one or more of such compounds, and methods of using the compounds to inhibit, regulate or modulate collagen formation or maturation as a therapeutic approach towards the treatment or prevention of diseases related to, or associated with, unregulated collagen production or maturation.
In one embodiment, compounds which are capable of inhibiting C-proteinase according to the invention, and which can therefore be used in methods to inhibit, modulate or regulate collagen production or maturation or to treat or prevent diseases related to, or associated with, unregulated collagen production or maturation are generally substituted hydantoins according to structural formulae (Ia) and (Iaxe2x80x2): 
or pharmaceutically acceptable salts thereof, wherein:
m is an integer from 0 to 3 when X is NR19 or an integer from 1 to 3 when X is S or O;
p is an integer from 1 to 4 when X is NR19 or an integer from 1 to 3 when X is S or O;
n is and integer from 1 to 3;
X is NR19, S or O;
R19 is selected from the group consisting of hydrogen, (C1-C5) alkyl and (C1-C5) substituted alkyl;
R1 and R2 are each independently selected from the group consisting of (C1-C10) alkyl, (C1-C10) substituted alkyl, (C3-C10) alkenyl, (C3-C10) substituted alkenyl, (C3-C10) alkynyl, (C3-C10) substituted alkynyl, (C3-C20) cycloalkyl, (C3-C20) substituted cycloalkyl, 3-20 membered heterocycloalkyl, 3-20 membered substituted heterocycloalkyl, (C5-C20) aryl, (C5-C20) aryl independently substituted with one or more Y1, alkaryl, alkaryl independently substituted with one or more Y1, 5-20 membered heteroaryl, 5-20 membered heteroaryl independently substituted with one or more Y1, 6-26 membered alk-heteroaryl, and 6-26 membered alk-heteroaryl independently substituted with one or more Y1;
R20 and R21 are each independently selected from the group consisting of hydrogen, (C1-C10) alkyl, (C6-C26) alkaryl, and 6-26 membered alk-heteroaryl; and
each Y1 is independently selected from the group consisting of -halogen, -trihalomethyl, xe2x80x94R, xe2x80x94C(O)OR, xe2x80x94CN, xe2x80x94C(O)xe2x80x94NRxe2x80x94OR, xe2x80x94C(NRR)xe2x95x90Nxe2x80x94OR, xe2x80x94C(O)xe2x80x94R, xe2x80x94C(O)NRR, xe2x80x94C(S)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94NRR, xe2x80x94NO2, xe2x80x94N3, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94OR, xe2x80x94P(O) (OH) (NRR), xe2x80x94P(O) (OH)2, xe2x80x94SO2R, xe2x80x94S(O) xe2x80x94R, xe2x80x94SO3H, xe2x80x94SR, xe2x80x94SO2xe2x80x94NRR, xe2x80x94OCF3 and -heteroaryl; where each R is independently selected from the group consisting of hydrogen, (C1-C8) alkyl, (C3-C8) alkenyl, (C5-C20) aryl, (C6-C26) alkaryl, 6-20 membered heteroaryl and 6-26-membered alk-heteroaryl;
with the proviso that the sum of m and p is 1, 2, 3 or 4.
In a preferred embodiment, the compounds of the present invention are compounds according to formula (Iaxe2x80x2) in which R20 and R21 are each hydrogen and R1, R2, and n are as defined for formulae (Ia) and (Iaxe2x80x2).
In another preferred embodiment, the compounds of the present invention are compounds according to formula (Ia) in which m and p are each 1. In this embodiment, X, R1, and R2 are as defined for formulae (Ia) and (Iaxe2x80x2).
In still another preferred embodiment, the compounds of the present invention are compounds according to formulae (Ia) and (Iaxe2x80x2) in which R1 is adamantyl. In this embodiment, X, R20, R21, R2, m, p, and n are as defined for formulae (Ia) and (Iaxe2x80x2).
In another preferred embodiment, the compounds of the present invention are compounds according to formulae (Ia) and (Iaxe2x80x2) in which R1 is cyclohexyl. In this embodiment, X, R20, R21, R2, m, p, and n are as defined for formulae (Ia) and (Iaxe2x80x2).
In a preferred embodiment, the compounds of the invention are generally substituted hydantoin compounds of the structural formula (Ib): 
or pharmaceutically acceptable salts thereof, wherein:
a is 2, 3, 4, or 5; and
R1, R2, and R20 are as defined for formula (Ia)
One group of preferred compounds according to formula (Ib) are those compounds having structural formula (II): 
or pharmaceutically acceptable salts thereof, wherein:
a is as defined for formula (Ib);
R1 and R20 are as defined as in formula (Ia); and
R3, R4, R5, R6, and R7 are each independently selected from the group consisting of -hydrogen, -halogen, -trihalomethyl, xe2x80x94R, xe2x80x94C(O)OR, xe2x80x94CN, xe2x80x94C(O)xe2x80x94NRxe2x80x94OR, xe2x80x94C(NRR)xe2x95x90Nxe2x80x94OR, xe2x80x94C(O)xe2x80x94R, xe2x80x94C(O)NRR, xe2x80x94C(S)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94NRR, xe2x80x94NO2, xe2x80x94N3, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O) xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O) xe2x80x94OR, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C (S) xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O) R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94OR, xe2x80x94P(O)(OH) (NRR), xe2x80x94P(O)(OH)2, xe2x80x94SO2R, xe2x80x94S(O) xe2x80x94R, xe2x80x94SO3H, xe2x80x94SR, xe2x80x94SO2xe2x80x94NRR, xe2x80x94OCF3 and -tetrazole; where each R is independently selected from the group consisting of xe2x80x94H, (C1-C8) alkyl, (C3-C8) alkenyl, (C3-C8) alkynyl, (C5-C20) aryl, (C6-C26) alkaryl, 6-20 membered heteroaryl and 6-26-membered alk-heteroaryl.
One group of preferred compounds according to formula (II) are compounds in which R20 is hydrogen.
Another group of preferred compounds according to formula (II) are those compounds wherein a is 3. Particularly preferred compounds according to this aspect of the invention are those compounds in which R3, R4, R5, R6, and R7 are each hydrogen. Particularly preferred compounds according to this aspect of the invention are compounds in which R20 is hydrogen and R1 is defined in Table 1. In Table 1, the connecting bond is dashed.
Another group of preferred compounds according to structure (Ib) are those compounds according to structural formula (III): 
or pharmaceutically acceptable salts thereof, wherein:
R8, R9 and R10 are each independently selected from the group consisting of -halogen, -trihalomethyl, xe2x80x94R, xe2x80x94C(O)OR, xe2x80x94CN, xe2x80x94C(O)xe2x80x94NRxe2x80x94OR, xe2x80x94C(NRR)xe2x95x90Nxe2x80x94OR, xe2x80x94C(O)xe2x80x94R, xe2x80x94C(O)NRR, xe2x80x94C(S)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94NRR, xe2x80x94NO2, xe2x80x94N3, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94OR, xe2x80x94P(O) (OH) (NRR), xe2x80x94P(O) (OH)2, xe2x80x94SO2R, xe2x80x94S(O)xe2x80x94R, xe2x80x94SO3H, xe2x80x94SR, xe2x80x94SO2xe2x80x94NRR, xe2x80x94OCF3 and -tetrazole; where each R is independently selected from the group consisting of xe2x80x94H, (C1-C8) alkyl, (C3-C8) alkenyl, (C3-C8) alkynyl, (C5-C20) aryl, (C6-C26) alkaryl, 6-20 membered heteroaryl, and 6-26-membered alk-heteroaryl; and
R20 is as defined for formula (Ia).
Particularly preferred compounds according to structure (III) are compounds in which R20 is xe2x80x94H, and R8, R9, and R10 are as defined in Table 2.
Another group of preferred compounds according to structure (Ib) are those compounds wherein R1 is adamantyl. A particularly preferred class of compounds according to this embodiment are compounds in which
a is 3;
R2 is selected from the group consisting of (C1-C10) alkyl, (C1-C10) substituted alkyl, (C3-C10) alkenyl, (C3-C10) substituted alkynyl, (C3-C10) alkynyl, (C3-C10) substituted alkynyl, (C3-C20) cycloalkyl, (C3-C20) substituted cycloalkyl, (C5-C20) aryl, (C5-C20) aryl independently substituted with one or more Y1, alkaryl, alkaryl independently substituted with one or more Y1; and Y1 is as defined in structural formula (Ia); and
R20 is as defined in formula (Ia)
Particularly preferred compounds in accordance with this class of compounds are compounds in which R20 is hydrogen and R2 is as defined in Table 3. In Table 3, the connecting bond in dashed.
Another group of preferred compounds of the present invention are compounds according to structural formula (IV): 
or pharmaceutically acceptable salts thereof, wherein:
R11, R12, R13, R14, and R15 are each independently selected from the group consisting of -halogen, -trihalomethyl, xe2x80x94R, xe2x80x94C(O)OR, xe2x80x94CN, xe2x80x94C(O)xe2x80x94NRxe2x80x94OR, xe2x80x94C(NRR)xe2x95x90Nxe2x80x94OR, xe2x80x94C(O)xe2x80x94R, xe2x80x94C(O)NRR, xe2x80x94C(S)NRR, xe2x80x94C(NRR)xe2x95x90NR, xe2x80x94NRR, xe2x80x94NO2, xe2x80x94N3, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94NRxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)xe2x80x94OR, xe2x80x94NRxe2x80x94SO2xe2x80x94R, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(O)R, xe2x80x94Nxe2x80x94C(O)xe2x80x94NRR, xe2x80x94NRxe2x80x94C(S)xe2x80x94NRR, xe2x80x94OR, xe2x80x94P(O)(OH)(NRR), xe2x80x94P(O)(OH)2, xe2x80x94SO2R, xe2x80x94S(O)xe2x80x94R, xe2x80x94SR, xe2x80x94SO2xe2x80x94NRR, xe2x80x94OCF3 and -tetrazole; where each R is independently selected from the group consisting of xe2x80x94H, (C1-C8) alkyl, (C3-C8) alkenyl, (C5-C20) aryl, (C6-C26) alkaryl, 6-20 membered heteroaryl and 6-26-membered alk-heteroaryl; and
R20 is as defined in formula (Ia).
Particularly preferred compounds according to formula (IV) are compounds in which R20 is xe2x80x94H, and R11, R12, R13, R14, and R15 as defined in Table 4:
A group of preferred compounds according to formula (IV) are those compounds in which R14 and R15 are hydrogen. Particularly preferred compounds according to this class of compounds are compounds in which R20 is hydrogen and R11, R12, and R13 are as in Table 5.
A group of preferred compounds according to formula (Ib) are those compounds in which R2 is selected from the group consisting of 5-20 membered heteroaryl, 5-20 membered heteroaryl independently substituted with one or more Y1, 3-20 membered heterocycloalkyl, 3-20 membered substituted heterocycloalkyl, 2-26 membered alk-heteroaryl, and 6-26 membered alk-heteroaryl independently substituted with one or more Y1; and
Y1 is as defined in formula (Ia).
Particularly preferred compounds according to this embodiment are compounds in which a is 3, R20 is xe2x80x94H, and R1 is adamantyl. Preferred compounds in this class include compounds of Table 6. In Table 6, the connecting bond is dashed.
Another group of preferred compounds according to structural formula (Ib) are those compounds wherein R1 is cyclohexyl. Particularly preferred compounds according to this aspect of the invention are those compounds in which a is 3. Even more preferred are compounds in this embodiment in which R20 is hydrogen and R2 is as defined in Table 7. In Table7, the connecting bond is dashed.
Yet another group of preferred compounds according to formula (Ia) are those compounds in which X is S. Particularly preferred compounds in which X is S are those compounds in which m is 1, p is 1 and R1 is adamantyl. Of the compounds of this class, compound 199 is particularly preferred: 
Another group of preferred compounds according to formula (Ia) are those compounds in which X is NR19. Particularly preferred compounds in this embodiment are those compounds in which m is 0, p is 3, and R1 is adamantyl. A particularly preferred compound in this embodiment is compound 200: 
Another group of preferred compounds according to formula (Ia) are those compounds in which X is O.
A group of preferred compounds according to formula (Iaxe2x80x2) are those compounds in which R1 is cyclohexyl, R20 and R21 are each methyl and R2 is as defined in Table 8. In Table 8, the connecting bond is dashed.
Another group of preferred compounds according to formula (Iaxe2x80x2) are those compounds in which R1 is adamantyl, R20 and R21 are each methyl and R2 is as defined in Table 9. In Table 9, the connecting bond is dashed.
The chemical structural formulae referred to herein may exhibit the phenomena of tautomerism, conformational isomerism, geometric isomerism and/or stereo isomerism. As the formulae drawings within this specification can only represent one of the possible tautomeric, conformational isomeric, geometric isomeric or stereo isomeric forms, it should be understood that the invention encompasses any tautomeric, conformational isomeric, geometric isomeric or stereo isomeric forms which exhibit biological or pharmacological activity as defined herein.
The compounds of the invention may be in the form of free acids, free bases or pharmaceutically effective acid addition or base addition salts. Such acid addition salts can be readily prepared by treating a compound with a pharmaceutically acceptable acid. Pharmaceutically acceptable acids include, by way of example and not limitation, inorganic acids such as hydrohalic acids (hydrochloric, hydrobromic, etc.), sulfuric acid, nitric acid, phosphoric acid, etc.; and organic acids such as acetic acid, propanoic acid, 2-hydroxyacetic acid, 2-hydroxypropanoic acid, 2-oxopropanoic acid, propandioic acid, butandioic acid, etc. Conversely, the acid addition salt can be converted into the free base form by treatment with alkali. Appropriate base addition salts can be readily prepared by treating a compound with a pharmaceutically acceptable base.
In addition to the above compounds and their pharmaceutically acceptable salts, the invention is further directed, where applicable, to solvated as well as unsolvated forms of the compounds (e.g. hydrated forms) exhibiting biological or pharmacological activity as defined herein.
The compounds of the invention may be prepared by any process known to be applicable to the preparation of chemical compounds. Suitable processes are well-known in the art. Preferred processes are illustrated by the representative examples. Necessary starting materials may be obtained commercially or by standard procedures of organic chemistry.
By way of example, the compounds of the invention can be conveniently prepared by schemes (I) thru (XII) below: 
In Scheme (I), compounds of formula (Ib) are synthesized and a, R1, R2, and R20 are as defined for formula (Ib).
According to scheme (I), D,L-2-amino-(C2-C5) alkanedioic acid 1 was mixed with SOCl2 in MeOH to yield D,L-2-amino-(C2-C5)alkanedioic acid dimethyl ester hydrochloride 2. To compound 2 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine to (0.55 mM), aldehyde 3 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 4 (0.55 mM) was added. The mixture was shaken for an additional twelve hours and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, and concentrated to yield residue 5. Residue 5 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 ml of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield compound of formula (Ib). Compound of formula (Ib) was triturated in ether/hexane (1.1, 5 mL), and dried in vaccuo.
D,L-2-amino-(C2-C5)alkanedioic acid compounds 1 in which R20 is hydrogen are commercially available. Compounds 1 in which R20 is a (C1-C10) alkyl, (C6-C26) alkaryl, and 6-26 membered alk-heteroaryl are synthesized by Strecker synthesis as follows: 
See, for example, Gal et al., 1954, J. Am. Chem. Soc. 76, 4181, Izumi et al., 1965, Bull. Chem. Soc. Jpn., 38, 1338, Weinges et al., 1971, Chem. Ber., 104, 3594. 
In Scheme (II), compounds of formula (II) are synthesized. Accordingly, R1, R3, R4, R5, R6, R7, R20 and a are as defined in formula (II). Compounds of Scheme (II) are synthesized according to Scheme (I) by using compound 10 for aldehyde 3. 
In Scheme (III), a, and R20 are as defined for formula (Ib). R2 is selected from the group consisting of (C1-C10) alkyl, (C1-C10) substituted alkyl, (C3-C10) alkenyl, (C3-C10) substituted alkenyl, (C3-C10) alkynyl, (C3-C10) substituted alkynyl, (C3-C20) cycloalkyl, (C3-C20) substituted cycloalkyl, (C5-C20) aryl, (C5-C20) aryl independently substituted with one or more Y1, alkaryl, alkaryl independently substituted with one or more Y1; where Y1 is as defined in structural formula (Ia). According to Scheme (III), D,L-2-amino-(C2-C5) alkanedioic acid 15 was mixed with SOCl2 in MeOH to yield D,L-2-amino-(C2-C5)alkanedioic acid dimethyl ester hydrochloride 16. To compound 16 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 17 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then adamantyl-isocynate 19 (0.55 mM) was added. The mixture was shaken for an additional twelve hours, and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, concentrated and dried to yield residue 20. Residue 20 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 ml of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield compound of formula (V). Compound of formula (V) was triturated in ether/hexane (1.1, 5 mL), and dried in vaccuo.
D,L-2-amino-(C2-C5) alkanedioic acid compounds 15 in which R20 is hydrogen are commercially available. Compounds 15 in which R20 is other than hydrogen may be prepared by Strecker synthesis as described in Scheme (I). 
In Scheme (IV), compounds of formula (IV) are synthesized. Accordingly, a, R11, R12, R13, R14, and R15 and R20 are as defined in formula (IV). Compounds are synthesized as described in Scheme (I) by using aldehyde 25 for compound 3 and Adamantyl-isocynate 27 for compound 4. 
In Scheme (V), a, R20, and R2 are as defined in formula (Ib). In Scheme (V), compounds 37 are synthesized using the synthesis described in Scheme (I), where cyclohexyl-isocynate 35 is used for compound 4. 
In Scheme (VI), R1, R2, m, and p are as defined in formula (Ia) and Fmoc is 9-fluorenylmethyl carbamate. Suitable amino protecting groups, G, may be found, for example, in Greene and Wuts, 1991, Protective Groups in Organic Synthesis, John Wiley and Sons, New York. In a preferred embodiment, G is tert-butoxycarbonyl.
According to Scheme (VI), compounds of formula (Ia) in which X is NH are synthesized. Compound 38 was mixed with piperidine to yield compound 39. To compound 39 (O.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 40 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 41 (0.55 mM) was added. The mixture was shaken for an additional twelve hours and then quenched with 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, and concentrated to yield intermediate. Intermediate, in which the protecting group G was tert butoxycarbonyl, was treated with trifluoroacetic acid in CH2Cl2 (5 percent, 5 mL) for five hours and then concentrated. The resulting residue was dissolved in 5 mL of CH2Cl2 and triethylamine and compound 42 were added. The mixture was stirred for twelve hours at room temperature. The reaction mixture was concentrated and purified by silica gel chromatography to give residue 43. Residue 43 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) 44 for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 mL of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield compound 45. 
In Scheme (VII), compounds of formula (Ia) in which X is NR19 are synthesized. Accordingly, m, p, R1, R2, and R19 are as defined in formula (Ia). According to Scheme (VII), compound 43, which was synthesized as detailed in Scheme (VI), was mixed with compound 52 to yield compound 53. Compound 53 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 mL of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield 54. 
In Scheme (VIII), compounds of formula (Ia) in which X is NH are synthesized. Accordingly, p, R1 and R2 are as defined in formula (Ia). According to Scheme (VIII), D,L-2-amino-(C2-C5) alkanedioic acid 55 is mixed with SOCl2 in MeOH to yield D,L-2-amino-(C2-C5)alkanedioic acid dimethyl ester hydrochloride 56. To compound 56 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 57 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 58 (0.55 mM) was added. The mixture was shaken for an additional twelve hours and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, concentrated and dried to yield residue 59. Residue 59 was treated with LiOH to yield compound 60. Compound 60 was mixed with diphenylphosphoryl azide, triethylamine, and ethylene glycol and refluxed at room temperature to form isocyanate 61. Isocyanate 61 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 mL of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield N-hydroxy-urea 62. 
In Scheme (IX), compounds of formula (Ia) in which X is O are synthesized. Accordingly, R1, R2, m, and p are as defined for formula (Ia) and Fmoc is 9-fluorenylmethyl carbamate. According to Scheme (IX), Compound 63 was combined with compound 64 in the presence of a base to yield compound 65. One suitable base was sodium hydride. Compound 65 was deprotected using piperidine to yield the corresponding free amine 66. Additionally, deprotection of the Fmoc group of compound 65 may be effected by the use of other amine bases as described in Greene and Wuts, supra, pages 318-319. To compound 66 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 67 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 68 (0.55 mM) is added. The mixture was shaken for an additional twelve hours and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO1, filtered, and concentrated to yield residue 69. Residue 69 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 ml of CH2Cl2. The organic phase was washed with water (5 mL) , dried over MgSO4, filtered and concentrated to yield residue 70. Residue 70 was triturated in ether/hexane (1.1, 5 mL), and dried in vaccuo. 
In Scheme (X), compounds of formula (Ia) in which X is S are synthesized. Accordingly, R1, R2, m and p are as defined for formula (Ia) and Fmoc is 9-fluorenylmethyl carbamate.
According to Scheme (X), compound 71 was combined with compound 72 in the presence of a base to yield compound 73. One suitable base was sodium hydride. Compound 73 was deprotected using piperidine to yield the corresponding free amine 74. Additionally, deprotection of the Fmoc group of compound 74 may be effected by the use of other amine bases as described in Greene and Wuts, supra, pages 318-319. To compound 74 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 75 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 76 (0.55 mM) was added. The mixture was shaken for an additional twelve hours and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, concentrated and dried to yield residue 77. Residue 77 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 ml of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield residue 78. Residue 78 was triturated in ether/hexane (1.1, 5 mL), and dried in vaccuo. 
In Scheme (XI), compounds of formula (Iaxe2x80x2) are synthesized. Accordingly, R1, R2, R20, R21, and n are as defined in formula (Iaxe2x80x2). According to scheme (XI), D,L-2-amino-(C2-C5)alkanedioic acid 80 is prepared by Strecker synthesis from compound 79. See, for example, Gal et al., 1954, J. Am. Chem. Soc. 76, 4181, Izumi et al., 1965, Bull. Chem. Soc. Jpn., 38, 1338, Weinges et al., 1971, Chem. Ber., 104, 3594. Compound 80 is mixed with SOCl2 in MeOH to yield D,L-2-amino-(C2-C5)alkanedioic acid dimethyl ester hydrochloride, the amino group of which was protected by tert-butylcarbamate (Boc) by reacting with Boc2O in the presence of triethylamine (TEA) and demethylaminopyridine (DMAP) to yield 81. Compound 81 was enolated by treating with lithium hexamethyldisilylamide [LiN(SiMe3)2], followed by the addition of R21Br to give compound 82. The Boc (tert-butylcarbonate) protecting group of compound 82 was removed by treating with three percent trifluoroacetic acid (TFA) in methylene chloride to furnish compound 83. To compound 83 (0.5 mM) in CH2Cl2 (5 mL) was added triethylamine (0.55 mM), aldehyde 84 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture was shaken at room temperature for six hours, and then isocynate 85 (0.55 mM) was added. The mixture was shaken for an additional twelve hours and then quenched with 1 mL of 1 N HCl. The organic phase was washed with water (5 mL), dried over MgSO4, filtered, and concentrated to yield residue 86. Residue 86 was treated with freshly prepared hydroxylamine solution (1.5 mL, 1.5 mM) for twelve hours, and then quenched with 2 mL of 1N HCl, and 6 ml of CH2Cl2. The organic phase was washed with water (5 mL), dried over MgSO4, filtered and concentrated to yield residue 87. Residue 87 was triturated in ether/hexane (1.1, 5 mL), and dried in vaccuo. 
In Scheme (XII), compounds of formula (Ia) in which X is NR19 and m is 0 are synthesized. Accordingly, R1, R2 and p are as defined in formula (I).
According to Scheme (XII), compound 88 is mixed with piperidine to yield compound 89. To compound 89 (0.5 mM) in CH2Cl2 (5 mL) is added triethylamine (0.55 mM), aldehyde 90 (0.55 mM), and sodium triacetoxyborohydride (0.7 mM). The reaction mixture is shaken at room temperature for 6 hours, and then isocyanate 91 (0.55 mM) is added. The mixture is shaken for additional 12 hours and the quenched with 1 N HCl. The organic phase is washed and dried to yield 92. The cyclization of compound 92 is carried out in the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene to afford 93. Tert-butoxycarbonyl (Boc) protecting group is removed upon treatment with 3% trifluoroacetic acid (TFA)/CH2Cl2 to provide compound 94, which, after being concentrated several times in ether, in turn reacts with 95 to yield compound 96. Compound 96 is then reacted with carbonyl diimidazole to give compound 97. Compound 97 is treated with freshly prepared hydroxylamine solution to yield compound 98.
An individual compound""s relevant activity and potency as an inhibitor of C-proteinase, to regulate or modulate collagen production or maturation and/or to treat disorders associated with unregulated collagen production may be determined using available techniques.
Typically, active compounds of the invention will inhibit 50% of the activity of C-proteinase at concentrations in the range of 100 micromolar (xcexcM) or less (i.e., those compounds exhibiting an IC50 of 100 xcexcM or less) using standard biochemical assays Dickson, 1953, Biochem. J. 55:170-171; Knight et al., 1992, FEBS 296:263-266. Those of skill in the art will appreciate that compounds exhibiting lower inhibitory concentrations (IC50s) are generally preferred for pharmacological applications; thus, preferably active compounds will exhibit IC50s that are less than 10 xcexcM, more preferably less than 1 xcexcM, even more preferably less than 100 nanomolar (nM) and even more preferably less than about 10 nM or 1 nM. However, as compounds which exhibit IC50s in the millimolar (mM) range can provide consequential pharmacological benefits, compounds which exhibit IC50s as high as 1 mM to 10 mM are considered to possess biological or pharmacological activity.
Alternatively, an in vitro procollagen assay may be used to determine the level of activity and effect of different compounds of the present invention on C-proteinase activity. In the procollagen assay, about 125 ug radiolabeled (14C) procollagen is added to 10 units/ml of chicken C-proteinase in a solution of 0.1 M Tris-HCl, 0.1 M NaCl, 0.02% Brij-35, and 5 mM CaCl2 in a total volume of 10 ul. The reaction is allowed to proceed for 15 minutes at 35xc2x0 C. and is stopped with one-half volume of 3xc3x97 stop/loading buffer (30 mM EDTA, 30% glycerol, 6% SDS, 0.006% Bromophenol-blue). Subsequently, the samples are heated to 100xc2x0 C. for 4 minutes, and resolved by SDS-PAGE (Novex) using 6% polyacryleamide gels. The protein bands are detected by autoradiography. The amount of enzyme activity is based on the disappearance of the band corresponding to uncleaved procollagen. The IC50 of inhibitors can be determined by plotting the percent activity versus inhibitor concentration and estimating the inhibitor concentration which results in 50% activity.
Disorders associated with unregulated collagen production or maturation can be treated with the compounds and compositions of the present invention. While not intending to be bound by any particular theory, it is believed that when administered to an animal subject, including a human, the compounds of the invention inhibit C-proteinase in vivo, thereby effectively modulating, regulating or inhibiting collagen production or maturation. As a consequence, the compounds are able to treat or prevent disorders associated with unregulated collagen production or maturation.
Collagen-related disorders which can be treated or prevented according to the invention include pathological fibrosis or scarring, such as endocardial sclerosis, idiopathic interstitial fibrosis, interstitial pulmonary fibrosis, perimuscular fibrosis, Symmers"" fibrosis, pericentral fibrosis, hepatitis, dermatofibroma, billary cirrhosis, alcoholic cirrhosis, acute pulmonary fibrosis, idiopathic pulmonary fibrosis, acute respiratory distress syndrome, kidney fibrosis/glomerulonephritis, kidney fibrosis/diabetic nephropathy, scleroderma/systemic, scleroderma/local, keloids, hypertrophic scars, severe joint adhesions/arthritis, myelofibrosis, corneal scarring, cystic fibrosis, muscular dystrophy (duchenne""s), cardiac fibrosis, muscular fibrosis/retinal separation, esophageal stricture, payronles disease. Further, fibrotic disorders may be induced or initiated by surgery such as scar revision/plastic surgeries, glaucoma, cataract fibrosis, corneal scarring, joint adhesions, graft vs. host disease, tendon surgery, nerve entrapment, dupuytren""s contracture, OB/GYN adhesions/fibrosis, pelvic adhesions, peridural fibrosis, restenosis. Still further fibrotic disorders may be induced by chemotherapy, including, for example lung fibrosis and the like.
The compounds described herein, or pharmaceutically acceptable addition salts or hydrates thereof, can be delivered to a subject, including a human, using a wide variety of routes or modes of administration. Suitable routes of administration include, but are not limited to, inhalation, transdermal, oral, rectal, transmucosal, intestinal and parenteral administration, including intramuscular, subcutaneous and intravenous injections.
The compounds described herein, or pharmaceutically acceptable salts and/or hydrates thereof, may be administered singly, in combination with other compounds of the invention, and/or in cocktails combined with other therapeutic agents. of course, the choice of therapeutic agents that can be co-administered with the compounds of the invention will depend, in part, on the condition being treated.
For example, the compounds of the invention can be administered in cocktails containing agents used to treat the pain and other symptoms and side effects commonly associated with fibrotic disorders. The compounds can also be administered in cocktails containing other agents that are commonly used to treat fibrotic disorders.
The active compound(s) may be administered per se or in the form of a pharmaceutical composition containing the active compound(s) and one or more pharmaceutically acceptable carriers, excipients or diluents. Administered compounds may be enantiomerically pure, or may be mixtures of enantiomers. Pharmaceutical compositions for use in accordance with the present invention may be formulated in conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen.
For injection, the agents of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks""s solution, Ringer""s solution, or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
For oral administration, the compounds can be formulated by combining the active compound(s) with pharmaceutically acceptable carriers well known in the art. Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient to be treated. Pharmaceutical preparations for oral use can be obtained solid excipient, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP). If desired, disintegrating agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.
Pharmaceutical preparations which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. All formulations for oral administration should be in dosages suitable for such administration.
For buccal administration, the compositions may take the form of tablets or lozenges formulated in conventional manner.
For administration by inhalation, the compounds for use according to the present invention are conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of e.g. gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch.
The compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
Pharmaceutical formulations for parenteral administration include aqueous solutions of the active compounds in water-soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.
Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.
In addition to the formulations previously described, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation or transcutaneous delivery (for example subcutaneously or intramuscularly), intramuscular injection or a transdermal patch. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
The pharmaceutical compositions also may comprise suitable solid or gel phase carriers or excipients. Examples of such carriers or excipients include but are not limited to calcium carbonate, calcium phosphate, various sugars, starches, cellulose derivatives, gelatin, and polymers such as polyethylene glycols.
5.5.1 Effective Dosage.
Pharmaceutical compositions suitable for use in the present invention include compositions wherein the active ingredients are contained in an effective amount to achieve its intended purpose. More specifically, a therapeutically effective amount means an amount effective to prevent development of or to alleviate the existing symptoms of the subject being treated. Determination of the effective amounts is well within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein.
For any compound used in the method of the invention, the therapeutically effective dose can be estimated initially from cell culture assays. For example, a dose can be formulated in animal models to achieve a circulating concentration range that includes the IC50. as determined in cell culture (i.e., the concentration of the test compound which achieves a half-maximal inhibition of the C-proteinase activity). Such information can be used to more accurately determine useful doses in humans.
A therapeutically effective dose refers to that amount of the compound that results in amelioration of symptoms or a prolongation of survival in a patient. Toxicity and therapeutic efficacy of such compounds can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio between LD50 and ED50. Compounds which exhibit high therapeutic indices are preferred. The data obtained from these cell culture assays and animal studies can be used in formulating a range of dosage for use in human. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient""s condition. See, e.g., Fingl et al., 1975, in xe2x80x9cThe Pharmacological Basis of Therapeuticsxe2x80x9d, Ch. 1 p.1.
Dosage amount and interval may be adjusted individually to provide plasma levels of the active moiety which are sufficient to maintain the C-proteinase inhibiting effects, or minimal effective concentration (MEC). The MEC will vary for each compound but can be estimated from in vitro data; for example, the concentration necessary to achieve 50-90% inhibition of the C-proteinase using the assays described herein. Dosages necessary to achieve the MEC will depend on individual characteristics and route of administration. However, HPLC assays or bioassays can be used to determine plasma concentrations.
Dosage intervals can also be determined using MEC value. Compounds should be administered using a regimen which maintains plasma levels above the MEC for 10-90% of the time, preferably between 30-90% and most preferably between 50-90%.
In cases of local administration or selective uptake, the effective local concentration of the drug may not be related to plasma concentration.
The amount of composition administered will, of course, be dependent on the subject being treated, on the subject""s weight, the severity of the affliction, the manner of administration and the judgment of the prescribing physician.
5.5.2 Packaging
The compositions may, if desired, be presented in a pack or dispenser device which may contain one or more unit dosage forms containing the active ingredient. The pack may, for example, comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration. Compositions comprising a compound of the invention formulated in a compatible pharmaceutical carrier may also be prepared, placed in an appropriate container, and labeled for treatment of an indicated condition. Suitable conditions indicated on the label may include treatment of arthritis or any other fibrotic disorder.